scholarly journals Comparative Genomics of Pneumocystis Species Suggests the Absence of Genes for myo-Inositol Synthesis and Reliance on Inositol Transport and Metabolism

mBio ◽  
2014 ◽  
Vol 5 (6) ◽  
Author(s):  
Aleksey Porollo ◽  
Thomas M. Sesterhenn ◽  
Margaret S. Collins ◽  
Jeffrey A. Welge ◽  
Melanie T. Cushion
Keyword(s):  
Fission Yeast ◽  
Fungal Infections ◽  
Inositol Phosphate ◽  
Pathogenic Fungi ◽  
Phosphate Metabolism ◽  
Transport Pathways ◽  
Content Type ◽  
Inositol Phosphate Metabolism ◽  
Genes Encoding

ABSTRACTIn the context of deciphering the metabolic strategies of the obligate pathogenic fungi in the genusPneumocystis, the genomes of three species (P. carinii,P. murina, andP. jirovecii) were compared among themselves and with the free-living, phylogenetically related fission yeast (Schizosaccharomyces pombe). The underrepresentation of amino acid metabolism pathways compared to those inS. pombe, as well as the incomplete steroid biosynthesis pathway, were confirmed forP. cariniiandP. jiroveciiand extended toP. murina. All threePneumocystisspecies showed overrepresentation of the inositol phosphate metabolism pathway compared to that in the fission yeast. In addition to those known inS. pombe, four genes, encoding inositol-polyphosphate multikinase (EC 2.7.1.151), inositol-pentakisphosphate 2-kinase (EC 2.7.1.158), phosphoinositide 5-phosphatase (EC 3.1.3.36), and inositol-1,4-bisphosphate 1-phosphatase (EC 3.1.3.57), were identified in the two rodentPneumocystisgenomes,P. cariniiandP. murina. TheP. jiroveciigenome appeared to contain three of these genes but lacked phosphoinositide 5-phosphatase. Notably, two genes encoding enzymes essential formyo-inositol synthesis, inositol-1-phosphate synthase (INO1) and inositol monophosphatase (INM1), were absent from all three genomes, suggesting thatPneumocystisspecies are inositol auxotrophs. In keeping with the need to acquire exogenous inositol, two genes with products homologous to fungal inositol transporters, ITR1 and ITR2, were identified inP. cariniiandP. murina, whileP. jiroveciicontained only the ITR1 homolog. The ITR and inositol metabolism genes inP. murinaandP. cariniiwere expressed during fulminant infection as determined by reverse transcriptase real-time PCR of cDNA from infected lung tissue. Supplementation ofin vitroculture with inositol yielded significant improvement of the viability ofP. cariniifor days 7 through 14.IMPORTANCEMicrobes in the genusPneumocystisare obligate pathogenic fungi that reside in mammalian lungs and causePneumocystispneumonia in hosts with weakened immune systems. These fungal infections are not responsive to standard antifungal therapy. A long-termin vitroculture system is not available for these fungi, impeding the study of their biology and genetics and new drug development. Given that all genomes of thePneumocystisspecies analyzed lack the genes for inositol synthesis and contain inositol transporters,Pneumocystisfungi, likeS. pombe, appear to be inositol auxotrophs. Inositol is important for the pathogenesis, virulence, and mating processes inCandida albicansandCryptococcus neoformans, suggesting similar importance within thePneumocystisspecies as well. This is the first report to (i) characterize genes in the inositol phosphate metabolism and transport pathways inPneumocystisspecies and (ii) identify inositol as a supplement that improved the viability ofP. cariniiinin vitroculture.

scholarly journals Targeting the Homoserine Dehydrogenase of Paracoccidioides Species for Treatment of Systemic Fungal Infections

2017 ◽  
Vol 61 (9) ◽  
Author(s):  
Mariane C. Bagatin ◽  
Arethusa L. Pimentel ◽  
Débora C. Biavatti ◽  
Ernani A. Basso ◽  
Erika S. Kioshima ◽  
...  
Keyword(s):  
Tertiary Structure ◽  
Fungal Infections ◽  
Paracoccidioides Brasiliensis ◽  
Pathogenic Fungi ◽  
Dynamics Simulation ◽  
Homoserine Dehydrogenase ◽  
Content Type ◽  
Zinc Database ◽  
Etiological Agents

ABSTRACT This work evaluated new potential inhibitors of the enzyme homoserine dehydrogenase (HSD) of Paracoccidioides brasiliensis, one of the etiological agents of paracoccidioidomycosis. The tertiary structure of the protein bonded to the analogue NAD, and l-homoserine was modeled by homology. The model with the best output was subjected to gradient minimization, redocking, and molecular dynamics simulation. Virtual screening simulations with 187,841 molecules purchasable from the Zinc database were performed. After the screenings, 14 molecules were selected and analyzed by the use of absorption, distribution, metabolism, excretion, and toxicity criteria, resulting in four compounds for in vitro assays. The molecules HS1 and HS2 were promising, exhibiting MICs of 64 and 32 μg · ml−1, respectively, for the Pb18 isolate of P. brasilensis, 64 μg · ml−1 for two isolates of P. lutzii, and also synergy with itraconazole. The application of these molecules to human-pathogenic fungi confirmed that the HSD enzyme may be used as a target for the development of drugs with specific action against paracoccidioidomycosis; moreover, these compounds may serve as leads in the design of new antifungals.

scholarly journals Nanoemulsion as an Effective Treatment against Human-Pathogenic Fungi

mSphere ◽  
2019 ◽  
Vol 4 (6) ◽  
Author(s):  
Alexis Garcia ◽  
Yong Yi Fan ◽  
Sandeep Vellanki ◽  
Eun Young Huh ◽  
DiFernando Vanegas ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Fungal Infections ◽  
Pathogenic Fungi ◽  
Antifungal Drugs ◽  
Solid Organ ◽  
Therapeutic Approaches ◽  
Content Type ◽  
New Therapeutic Approaches ◽  
Development Of Resistance

ABSTRACT Infections triggered by pathogenic fungi cause a serious threat to the public health care system. In particular, an increase of antifungal drug-resistant fungi has resulted in difficulty in treatment. A limited variety of antifungal drugs available to treat patients has left us in a situation where we need to develop new therapeutic approaches that are less prone to development of resistance by pathogenic fungi. In this study, we demonstrate the efficacy of the nanoemulsion NB-201, which utilizes the surfactant benzalkonium chloride, against human-pathogenic fungi. We found that NB-201 exhibited in vitro activity against Candida albicans, including both planktonic growth and biofilms. Furthermore, treatments with NB-201 significantly reduced the fungal burden at the infection site and presented an enhanced healing process after subcutaneous infections by multidrug-resistant C. albicans in a murine host system. NB-201 also exhibited in vitro growth inhibition activity against other fungal pathogens, including Cryptococcus spp., Aspergillus fumigatus, and Mucorales. Due to the nature of the activity of this nanoemulsion, there is a minimized chance of drug resistance developing, presenting a novel treatment to control fungal wound or skin infections. IMPORTANCE Advances in medicine have resulted in the discovery and implementation of treatments for human disease. While these recent advances have been beneficial, procedures such as solid-organ transplants and cancer treatments have left many patients in an immunocompromised state. Furthermore, the emergence of immunocompromising diseases such as HIV/AIDS or other immunosuppressive medical conditions have opened an opportunity for fungal infections to afflict patients globally. The development of drug resistance in human-pathogenic fungi and the limited array of antifungal drugs has left us in a scenario where we need to develop new therapeutic approaches to treat fungal infections that are less prone to the development of resistance by pathogenic fungi. The significance of our work lies in utilizing a novel nanoemulsion formulation to treat topical fungal infections while minimizing risks of drug resistance development.

scholarly journals Basal Body Structures Differentially Affect Transcription of RpoN- and FliA-Dependent Flagellar Genes in Helicobacter pylori

2015 ◽  
Vol 197 (11) ◽  
pp. 1921-1930 ◽  
Author(s):  
Jennifer Tsang ◽  
Timothy R. Hoover
Keyword(s):  
Helicobacter Pylori ◽  
Basal Body ◽  
Protein Interactions ◽  
Transcript Levels ◽  
Content Type ◽  
Genes Encoding ◽  
H Pylori ◽  
Flagellar Biogenesis ◽  
Flagellar Genes

ABSTRACTFlagellar biogenesis inHelicobacter pyloriis regulated by a transcriptional hierarchy governed by three sigma factors, RpoD (σ80), RpoN (σ54), and FliA (σ28), that temporally coordinates gene expression with the assembly of the flagellum. Previous studies showed that loss of flagellar protein export apparatus components inhibits transcription of flagellar genes. The FlgS/FlgR two-component system activates transcription of RpoN-dependent genes though an unknown mechanism. To understand better the extent to which flagellar gene regulation is coupled to flagellar assembly, we disrupted flagellar biogenesis at various points and determined how these mutations affected transcription of RpoN-dependent (flaBandflgE) and FliA-dependent (flaA) genes. The MS ring (encoded byfliF) is one of the earliest flagellar structures assembled. Deletion offliFresulted in the elimination of RpoN-dependent transcripts and an ∼4-fold decrease inflaAtranscript levels. FliH is a cytoplasmic protein that functions with the C ring protein FliN to shuttle substrates to the export apparatus. Deletions offliHand genes encoding C ring components (fliMandfliY) decreased transcript levels offlaBandflgEbut had little or no effect on transcript levels offlaA. Transcript levels offlaBandflgEwere elevated in mutants where genes encoding rod proteins (fliEandflgBC) were deleted, while transcript levels offlaAwas reduced ∼2-fold in both mutants. We propose that FlgS responds to an assembly checkpoint associated with the export apparatus and that FliH and one or more C ring component assist FlgS in engaging this flagellar structure.IMPORTANCEThe mechanisms used by bacteria to couple transcription of flagellar genes with assembly of the flagellum are poorly understood. The results from this study identified components of theH. pyloriflagellar basal body that either positively or negatively affect expression of RpoN-dependent flagellar genes. Some of these basal body proteins may interact directly with regulatory proteins that control transcription of theH. pyloriRpoN regulon, a hypothesis that can be tested by examining protein-protein interactionsin vitro.

Differential induction of inositol phosphate metabolism by three adipokinetic hormones

1997 ◽  
Vol 130 (1-2) ◽  
pp. 131-139 ◽  
Author(s):  
Simon F Vroemen ◽  
Wil J.A Van Marrewijk ◽  
Jeroen De Meijer ◽  
Aloys Th.M Van den Broek ◽  
Dick J Van der Horst
Keyword(s):  
Inositol Phosphate ◽  
Phosphate Metabolism ◽  
Inositol Phosphate Metabolism ◽  
Differential Induction

scholarly journals Antimicrobial Activity of Ceftazidime-Avibactam Tested against Multidrug-Resistant Enterobacteriaceae and Pseudomonas aeruginosa Isolates from U.S. Medical Centers, 2013 to 2016

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Helio S. Sader ◽  
Mariana Castanheira ◽  
Dee Shortridge ◽  
Rodrigo E. Mendes ◽  
Robert K. Flamm
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Multidrug Resistant ◽  
Drug Resistant ◽  
Large Collection ◽  
Content Type ◽  
Negative Results ◽  
Medical Centers ◽  
Extensively Drug Resistant ◽  
Genes Encoding

ABSTRACT The in vitro activity of ceftazidime-avibactam and many comparator agents was determined against various resistant subsets of organisms selected among 36,380 Enterobacteriaceae and 7,868 Pseudomonas aeruginosa isolates. The isolates were consecutively collected from 94 U.S. hospitals, and all isolates were tested for susceptibility by reference broth microdilution methods in a central monitoring laboratory (JMI Laboratories). Enterobacteriaceae isolates resistant to carbapenems (CRE) and/or ceftazidime-avibactam (MIC ≥ 16 μg/ml) were evaluated for the presence of genes encoding extended-spectrum β-lactamases and carbapenemases. Ceftazidime-avibactam inhibited >99.9% of all Enterobacteriaceae at the susceptible breakpoint of ≤8 μg/ml and was active against multidrug-resistant (MDR; n = 2,953; MIC50/90, 0.25/1 μg/ml; 99.2% susceptible), extensively drug-resistant (XDR; n = 448; MIC50/90, 0.5/2 μg/ml; 97.8% susceptible), and CRE (n = 513; MIC50/90, 0.5/2 μg/ml; 97.5% susceptible) isolates. Only 82.2% of MDR Enterobacteriaceae (n = 2,953) and 64.2% of ceftriaxone-nonsusceptible Klebsiella pneumoniae (n = 1,063) isolates were meropenem susceptible. Among Enterobacter cloacae (22.2% ceftazidime nonsusceptible), 99.8% of the isolates, including 99.3% of the ceftazidime-nonsusceptible isolates, were ceftazidime-avibactam susceptible. Only 23 of 36,380 Enterobacteriaceae (0.06%) isolates were ceftazidime-avibactam nonsusceptible, including 9 metallo-β-lactamase producers and 2 KPC-producing strains with porin alteration; the remaining 12 strains showed negative results for all β-lactamases tested. Ceftazidime-avibactam showed potent activity against P. aeruginosa (MIC50/90, 2/4 μg/ml; 97.1% susceptible), including MDR (MIC50/90, 4/16 μg/ml; 86.5% susceptible) isolates, and inhibited 71.8% of isolates nonsusceptible to meropenem, piperacillin-tazobactam, and ceftazidime (n = 628). In summary, ceftazidime-avibactam demonstrated potent activity against a large collection (n = 44,248) of contemporary Gram-negative bacilli isolated from U.S. patients, including organisms resistant to most currently available agents, such as CRE and meropenem-nonsusceptible P. aeruginosa.

scholarly journals Analysis of Inositol Phosphate Metabolism by Capillary Electrophoresis Electrospray Ionization Mass Spectrometry (CE-ESI-MS)

2020 ◽  
Author(s):  
Danye Qiu ◽  
Miranda S. Wilson ◽  
Verena B. Eisenbeis ◽  
Robert K. Harmel ◽  
Esther Riemer ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Capillary Electrophoresis ◽  
Electrospray Ionization ◽  
Electrospray Ionization Mass Spectrometry ◽  
Inositol Phosphate ◽  
Ionization Mass Spectrometry ◽  
Ionization Mass ◽  
Phosphate Metabolism ◽  
Esi Ms ◽  
Inositol Phosphate Metabolism

AbstractThe analysis of myo-inositol phosphates (InsPs) and myo-inositol pyrophosphates (PP-InsPs) is a daunting challenge due to the large number of possible isomers, the absence of a chromophore, the high charge density, the low abundance, and the instability of the esters and anhydrides. Given their importance in biology, an analytical approach to follow and understand this complex signaling hub is highly desirable. Here, capillary electrophoresis (CE) coupled to electrospray ionization mass spectrometry (ESI-MS) is implemented to analyze complex mixtures of InsPs and PP-InsPs with high sensitivity. Stable isotope labeled (SIL) internal standards allow for matrix-independent quantitative assignment. The method is validated in wild-type and knockout mammalian cell lines and in model organisms. SIL-CE-ESI-MS enables for the first time the accurate monitoring of InsPs and PP-InsPs arising from compartmentalized cellular synthesis pathways, by feeding cells with either [13C6]-myo-inositol or [13C6]-D-glucose. In doing so, we uncover that there must be unknown inositol synthesis pathways in mammals, highlighting the unique potential of this method to dissect inositol phosphate metabolism and signalling.

scholarly journals Identification and Predictions Regarding the Biosynthesis Pathway of Polyene Macrolides Produced by Streptomyces roseoflavus Men-myco-93-63

2021 ◽  
Vol 87 (10) ◽  
Author(s):  
Xing Han ◽  
Jiao Wang ◽  
Lianna Liu ◽  
Fengying Shen ◽  
Qingfang Meng ◽  
...  
Keyword(s):  
Fermentation Broth ◽  
Pathogenic Fungi ◽  
Biosynthetic Gene Cluster ◽  
Inhibitory Effect ◽  
Plant Diseases ◽  
Active Metabolites ◽  
Content Type ◽  
Polyene Macrolides ◽  
Alternative Agent

ABSTRACT A group of polyene macrolides mainly composed of two constituents was isolated from the fermentation broth of Streptomyces roseoflavus Men-myco-93-63, which was isolated from soil where potato scabs were repressed naturally. One of these macrolides was roflamycoin, which was first reported in 1968, and the other was a novel compound named Men-myco-A, which had one methylene unit more than roflamycoin. Together, they were designated RM. This group of antibiotics exhibited broad-spectrum antifungal activities in vitro against 17 plant-pathogenic fungi, with 50% effective concentrations (EC50) of 2.05 to 7.09 μg/ml and 90% effective concentrations (EC90) of 4.32 to 54.45 μg/ml, which indicates their potential use in plant disease control. Furthermore, their biosynthetic gene cluster was identified, and the associated biosynthetic assembly line was proposed based on a module and domain analysis of polyketide synthases (PKSs), supported by findings from gene inactivation experiments. IMPORTANCE Streptomyces roseoflavus Men-myco-93-63 is a biocontrol strain that has been studied in our laboratory for many years and exhibits a good inhibitory effect in many crop diseases. Therefore, the identification of antimicrobial metabolites is necessary and our main objective. In this work, chemical, bioinformatic, and molecular biological methods were combined to identify the structures and biosynthesis of the active metabolites. This work provides a new alternative agent for the biological control of plant diseases and is helpful for improving both the properties and yield of the antibiotics via genetic engineering.

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